Abstract
BackgroundMicrosatellite markers have proven useful in genetic studies in many organisms, yet microsatellite-based studies of the dengue and yellow fever vector mosquito Aedes aegypti have been limited by the number of assayable and polymorphic loci available, despite multiple independent efforts to identify them. Here we present strategies for efficient identification and development of useful microsatellites with broad coverage across the Aedes aegypti genome, development of multiplex-ready PCR groups of microsatellite loci, and validation of their utility for population analysis with field collections from Haiti.ResultsFrom 79 putative microsatellite loci representing 31 motifs identified in 42 whole genome sequence supercontig assemblies in the Aedes aegypti genome, 33 microsatellites providing genome-wide coverage amplified as single copy sequences in four lab strains, with a range of 2-6 alleles per locus. The tri-nucleotide motifs represented the majority (51%) of the polymorphic single copy loci, and none of these was located within a putative open reading frame. Seven groups of 4-5 microsatellite loci each were developed for multiplex-ready PCR. Four multiplex-ready groups were used to investigate population genetics of Aedes aegypti populations sampled in Haiti. Of the 23 loci represented in these groups, 20 were polymorphic with a range of 3-24 alleles per locus (mean = 8.75). Allelic polymorphic information content varied from 0.171 to 0.867 (mean = 0.545). Most loci met Hardy-Weinberg expectations across populations and pairwise FST comparisons identified significant genetic differentiation between some populations. No evidence for genetic isolation by distance was observed.ConclusionDespite limited success in previous reports, we demonstrate that the Aedes aegypti genome is well-populated with single copy, polymorphic microsatellite loci that can be uncovered using the strategy developed here for rapid and efficient screening of genome supercontig assemblies. These loci are suitable for genetic and population studies using multiplex-PCR.
Highlights
Microsatellite markers have proven useful in genetic studies in many organisms, yet microsatellite-based studies of the dengue and yellow fever vector mosquito Aedes aegypti have been limited by the number of assayable and polymorphic loci available, despite multiple independent efforts to identify them
Detailed genetic studies have emerged from linkage maps for Ae. aegypti generated from isozyme and mutant marker loci [2], RAPDs [3], RFLPs [4,5], and SSCPs [6]
Microsatellite identification, assays and utility Tandem Repeats Finder (TRF) [23] was used to systematically screen 42 wgs supercontig sequence assemblies in the Ae. aegypti genome for polymorphic single copy microsatellites (Figure 1)
Summary
Microsatellite markers have proven useful in genetic studies in many organisms, yet microsatellite-based studies of the dengue and yellow fever vector mosquito Aedes aegypti have been limited by the number of assayable and polymorphic loci available, despite multiple independent efforts to identify them. Of African origin, Ae. aegypti has successfully colonized most subtropical and tropical regions of the world, largely as a consequence of human activities This mosquito has been and remains the most commonly studied mosquito species, for genetic analyses of disease vector/pathogen interactions because it breeds in small water-holding containers, its eggs are resistant to desiccation and persist in a pre-embryonated state, and it readily adapts to laboratory culture. Availability of a partial Ae. aegypti genome sequence in 2005 provided the opportunity to perform genome scans for microsatellites and, an additional 13 polymorphic microsatellites were uncovered [21]
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